Mutant LRRK2(R1441G) BAC transgenic mice recapitulate cardinal features of Parkinson's disease

Abstract

Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common genetic cause of Parkinson's disease. We created a LRRK2 transgenic mouse model that recapitulates cardinal features of the disease: an age-dependent and levodopa-responsive slowness of movement associated with diminished dopamine release and axonal pathology of nigrostriatal dopaminergic projection. These mice provide a valid model of Parkinson's disease and are a resource for the investigation of pathogenesis and therapeutics.

Figure 1

LRRK2^R1441G BAC mice develop age-dependent motor deficits that responds to levodopa and apomorphine. (a) An age-dependent decline in rearing was detected from 3 to 6 months (wild-type nontransgenic littermates (WT), n = 5; LRRK2^R1441G BAC mice (TG) n = 5; P = 0.005). At 10–12 months, LRRK2^R1441G BAC mice had more significant reductions in rearing (WT, n = 14; TG, n = 17; P = 0.001). (b) This robust deficit in LRRK2^R1441G BAC mice was rescued by 20 mg per kg of body weight of levodopa with 6.25 mg per kg of benserazide but not by 2 mg per kg of levodopa. A similar rescue was achieved by 10 mg per kg of apomorphine. Data are presented as mean ± s.e.m. Asterisks indicate statistically significant differences (P < 0.05).

Figure 2

Neurochemical deficits in LRRK2^R1441G mice. In the presence of nomifensine, striatal dopamine release was significantly decreased in freely moving LRRK2^R1441G BAC mice, as seen by microdialysate analysis (F_1,16 = 5.7, P = 0.03, two-way ANOVA). Total spontaneous dopamine release, calculated as the area under the curve above baseline, was significantly decreased (35%, P = 0.03, two way ANOVA and Mann-Whitney test). [DA], dopamine concentration.

Figure 3

Morphologic abnormalities of mesencephalic dopamine neurons and their axons in LRRK2^R1441G BAC transgenic mice. (a) Tyrosine hydroxylase immunostaining of ventral mesencephalon. ML, medial lemniscus; MT, medial terminal nucleus. Rectangles in the upper panels are shown at a higher magnification in the lower panels. LRRK2^R1441G BAC transgenic mice showed a loss of tyrosine hydroxylase–positive dendrites in substantia nigra pars reticulata and a reduction of tyrosine hydroxylase neuron size in SNpc. (b) Normal optical density of tyrosine hydroxylase immunostaining of the whole striatum and dorsal-lateral quadrant. (c) In tyrosine hydroxylase–positive axons in the striatum and the ventral pallidum of LRRK2^R1441G BAC transgenic mice, we observed discrete foci of thickening with a beaded appearance (blue arrowheads) and fragmentation, large tyrosine hydroxylase–positive spheroid-like structures (blue arrow in the left panel and the insert in the upper right panel), and dystrophic neurites and enlarged axonal endings (lower right). The dystrophic neurites were found in all of the five LRRK2^R1441G mice (2.8 ± 0.6, s.e.m.) but were rarely seen in two of the seven nontransgenic littermates (0.4 ± 0.3; P = 0.003, t test). Dystrophic neurites were not found in the two WT-OX mice. Thus, expressed as number of neurites per area of the ventral pallidum (mm²), dystrophic neurites were 8.5-fold more prevalent among the transgenic mice than the controls. (d) In the dorsal striatum and piriform cortex, immunohistochemistry with AT8 revealed abnormal axon terminal enlargements and dystrophic neurites (arrows, top two left panels). EC, external capsule. AT8-positive structures are shown at a higher magnification in the right panels. A total of 42 AT8-positive dystrophic neurites were identified in the two LRRK2^R1441G mice that we examined. A single example was identified under blind conditions in one of the four nontransgenic mice, and none were found in the two WT-OX mice. Expressed as the number of neurites per forebrain coronal hemisection, 21 AT8-positive neurites were observed among the LRRK2^R1441G transgenics and only 0.17 among the control brains, a 124-fold difference. (e) Phosphorylated tau was markedly increased in the LRRK2^R1441G BAC transgenic mouse brains. Full-size western blots are presented in Supplementary Figure 7 online.